Methods of expanding t cells

Abstract

Provided herein are methods in the field of cell culture, specifically of culture and expansion of immune cells, e.g., T lymphocytes.

What is claimed is: 
     
         1 . A method of inducing a population of immune cells to proliferate, comprising contacting the immune cells with a ligand of a T cell receptor (TCR)/CD3 complex on said immune cells, and a ligand of a costimulatory molecule on said immune cells, such that said immune cells are caused to proliferate, wherein said ligands are comprised within a composition, wherein said composition comprises a liquid phase or multi-construct phase, and wherein said ligands are not immobilized on the surface of said composition. 
     
     
         2 . The method of  claim 1 , wherein said composition comprises a hydrogel, an albumin, or a hydrophobic molecule. 
     
     
         3 . The method of  claim 1 , wherein said composition is in the form of a bead. 
     
     
         4 . The method of  claim 1 , wherein said composition is substantially spherical. 
     
     
         5 . The method of  claim 1 , wherein said ligand of said TCR/CD3 complex is a macromolecular polypeptide binding agent. 
     
     
         6 . The method of  claim 5 , wherein said ligand is an antibody against CD3 (anti-CD3). 
     
     
         7 . The method of  claim 6 , wherein said antibody is polyclonal. 
     
     
         8 . The method of  claim 6 , wherein said antibody is monoclonal. 
     
     
         9 . The method of  claim 6 , wherein said antibody is the monoclonal antibody is OKT3 or G19-4 or a CD3-binding portion thereof. 
     
     
         10 . The method of  claim 1 , wherein said ligand of a costimulatory molecule is a ligand for CD28. 
     
     
         11 . The method of  claim 1 , wherein said ligand of said costimulatory molecule is a macromolecular polypeptide binding agent against CD28. 
     
     
         12 . The method of  claim 1 , wherein said ligand of said costimulatory molecule is an antibody against CD28 (anti-CD28). 
     
     
         13 . The method of  claim 12 , wherein said antibody is polyclonal. 
     
     
         14 . The method of  claim 12 , wherein said antibody is monoclonal. 
     
     
         15 . The method of  claim 1 , wherein said ligand of said costimulatory molecule is an antibody against CD9. 
     
     
         16 . The method of  claim 15 , wherein said antibody is polyclonal. 
     
     
         17 . The method of  claim 15 , wherein said antibody is monoclonal. 
     
     
         18 . The method of  claim 15 , wherein said antibody is ES5.2D8. 
     
     
         19 . The method of  claim 1 , wherein said ligand of said costimulatory molecule is B7-1 or B7-2 or a CD28-binding fragment thereof. 
     
     
         20 . The method of  claim 1 , wherein said composition comprises a hydrogel. 
     
     
         21 . The method of  claim 20 , wherein said hydrogel is a self-assembling hydrogel. 
     
     
         22 . The method of  claim 20 , wherein said hydrogel comprises a polyglutamate chain and vitamin E. 
     
     
         23 . The method of any of  claims 20 - 22  wherein said ligands are contained within a mesh of said hydrogel. 
     
     
         24 . The method of  claim 1 , wherein the composition comprises an oil, lipid or fluorocarbon, wherein said oil, lipid or fluorocarbon is a liquid. 
     
     
         25 . The method of  claim 24 , wherein said oil, lipid or fluorocarbon is present in the composition in the form of nanometer- or micrometer-scale droplets. 
     
     
         26 . The method of  claim 24 , wherein said ligands are present in said composition on the surface of said microdroplets. 
     
     
         27 . The method of  claim 26 , wherein said ligands are anti-CD3 and anti-CD28. 
     
     
         28 . The method of  claim 1 , wherein the composition comprises albumin. 
     
     
         29 . The method of  claim 28 , wherein said albumin is complexed with a liquid hydrophobic molecule. 
     
     
         30 . The method of  claim 29 , wherein said hydrophobic molecule is an oil, a lipid, or a fluorocarbon. 
     
     
         31 . The method of  claim 29  or  claim 30 , wherein said albumin is present on the surface of a droplet of said hydrophobic molecule. 
     
     
         32 . The method of  claim 29  or  claim 30 , wherein said albumin and said ligands are present on the surface of a droplet of said hydrophobic molecule. 
     
     
         33 . The method of any of  claims 28 - 32 , wherein the albumin is non-native albumin. 
     
     
         34 . The method of any of  claims 28 - 32 , wherein said albumin is denatured. 
     
     
         35 . The method of  claim 1 , wherein at least one of said ligands is conjugated to a hydrophobic molecule. 
     
     
         36 . The method of  claim 1 , wherein the CD3 ligand is conjugated to a hydrophobic molecule. 
     
     
         37 . The method of  claim 1 , wherein said ligand to a costimulatory molecule is conjugated to a hydrophobic molecule. 
     
     
         38 . The method of  claim 1 , wherein the ligand is a ligand of CD28. 
     
     
         39 . The method of  claim 1 , wherein each of said ligands is respectively conjugated to a hydrophobic molecule. 
     
     
         40 . The method of  claim 39 , wherein said ligands are anti-CD28 and anti-CD3. 
     
     
         41 . The method of  claim 39 , wherein said ligands are anti-CD3 and one or B7-1 or B7-2. 
     
     
         42 . The method of  claim 1 , wherein said composition comprises a hydrophobic molecule, wherein neither of said ligands is conjugated to said hydrophobic molecule. 
     
     
         43 . The method of  claim 39 , wherein said composition comprises a hydrophobic molecule, wherein said ligands are anti-CD28 and anti-CD3, and neither of said anti-CD28 or said anti-CD3 is conjugated to said hydrophobic molecule. 
     
     
         44 . The method of  claim 39 , wherein said composition comprises a hydrophobic molecule, wherein said ligands are anti-CD3 and one of B7-1 or B7-2, and neither of said anti-CD3 or said one of B7-1 or B7-2 is conjugated to said hydrophobic molecule. 
     
     
         45 . The method of any of  claims 35 - 44 , wherein said hydrophobic molecule self-aggregates in aqueous solution to form a micelle. 
     
     
         46 . The method of any of  claims 35 - 44 , wherein said hydrophobic molecule is a lipid. 
     
     
         47 . The method of any of  claims 35 - 44 , wherein said hydrophobic molecule is an aliphatic chain. 
     
     
         48 . The method of any of  claims 35 - 44 , wherein said hydrophobic molecule is a fluorocarbon. 
     
     
         49 . The method of any of  claims 2 - 48 , wherein said albumin is human serum albumin. 
     
     
         50 . The method of any of  claims 1 - 49 , wherein said immune cells are T cells. 
     
     
         51 . The method of  claim 50 , wherein said T cells are CD4+ T cells. 
     
     
         52 . The method of  claim 50 , wherein said T cells are CD8+ T cells. 
     
     
         53 . The method of  claim 50 , wherein said T cells are Treg cells. 
     
     
         54 . The method of any of  claims 1 - 53 , wherein the immune cells are expanded about 100-fold to about 100,000-fold. 
     
     
         55 . The method of any of  claims 1 - 54 , wherein the composition comprises a liquid phase or multi-phase system that is largely immiscible with aqueous media between 2° C. and 44° C. 
     
     
         56 . The method of  claim 1 , wherein the composition presents as a colloid or an emulsion with aqueous media between 2° C. and 44° C. upon homogenization. 
     
     
         57 . The method of any of  claims 1 - 56 , additionally comprising contacting the immune cells with interleukin-2 (IL-2). 
     
     
         58 . The method of any of  claims 1 - 57 , additionally comprising contacting the immune cells with a phorbol ester.